The mechanisms that dictate the final program of gene expression in a fully differentiated cell can be revealed by starting at either end of the regulatory cascade. To examine the series of controls operating on cells of the oligodendrocyte lineage, we have begun with one of the final targets of regulation in myelinating glial cells, proteolipid protein (PLP). Expression libraries were screened with DNA probes corresponding to PLP cis ~regulatory elements by a method that relies on the detection of DNA~protein interactions five novel clones (named MyTI~V for Myelin Transcription Factor) were isolated. The most extensively characterized clone of this screen, MyTI, is highly expressed in the developing nervous system, primarily in the nuclei of progenitor cells. When progenitors are induced to differentiate into oligodendrocytes in vitro, the nuclear form of MyTI disappears and the cells transiently express MyTI in the cytoplasm. The consensus binding site for MyTI is represented in the PLP promoter as well as in other myelin gene promoters, presenting a mode of coordinate control for these genes during oligodendrocyte development. The isolation of clones encoding transcriptional regulatory proteins permits a search for the growth factors and other molecules that are critical to the initiation and maintenance of myelin gene transcription during development and regeneration. In a spinal cord contusion model, we have found that the putative myelin transcription factor, MyTI, is dramatically up~regulated following injury. The expression of MyTI precedes the induction of myelin expression and therefore provides a handle for examining the molecular events underlying the remyelinating state. Mutations in the major myelin protein PLP result in a devastating loss of white matter in the X~ linked disease of man (Pelizaeus~Merzbacher disease) and animals. To investigate whether the PLP locus has multiple roles in myelinating cells that would explain the pleiotropic phenotypes observed in the PLP mutants, we generated transgenic mice which express either PLP or its alternatively spliced isoform, DM20. Neither the PLP transgene nor the DM20 transgene alone restored myelin expression in mice. Only a combination of the two transgenes substantially increased myelination, suggesting that the two alternatively spliced products of the PLP locus perform distinct functions in oligodendrocytes. Thus, the transgenic approach offers a suitable in vivo system for dissecting gene function, and will continue to be applied to our studies of other genes in the nervous system.